Scanning Microscopy


Ultra-small gold colloids bound to immunolabeled ultrathin resin sections were visualized using transmission, scanning, and scanning transmission electron microscopy (TEM, SEM, STEM). The best marker contrast is obtained in a field emission STEM (200 kV) equipped with a high-angle annular dark-field (HAADF) detector. HAADF STEM renders possible the simultaneous visualization of ultra-small gold and ultrastructural details in unstained resin sections, and an overall presentation of a labeled E. coli cell.

For routine work, an enhancement step is a prerequisite for easy detection of bound marker molecules. Five different silver enhancing solutions were tested for their suitability for ultra-small gold intensification. Enhancers lacking the protective colloid gum arabic exhibit lower quality with regard to efficiency and homogeneity of enhancement. This problem can be overcome by adding gum arabic. Silver enhancement generally results in heterogeneously sized particles. This is most probably due to the heterogeneous original gold colloid probe. In general, an estimation of enhancement efficiency is associated with difficulties depending on experimental conditions and the electron microscopic imaging modes used. Only a low number of the ultra-small gold particles seems to remain unenhanced or poorly enhanced when treated with "high-quality" enhancers. On-section labeling of ultrathin resin sections with silver-enhanced ultra-small gold markers also offers the possibility of high-resolution immunolabeling experiments at the light microscopic level.

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